Process for forming a lignin concentrate



STATESPATENT OFFICE PROCESS FOR FORMING A LIGNIN CONCENTRATE ration of New'Jersey No Drawing. Application December 3, 1948, Serial No. 63.452

5 Claims.

This invention relates to an improved process for enriching the lignin content of wood, and more especially it deals with such a process involving the action of certain specified microbic organisms under certain specified conditions.

It is known that wood is attacked by many microorganisms. Some microorganisms, e. g. certain Basidiomycetes of the families Agaricaceae, Polyporaceae, etc., attack or break down the cellulose of wood. Certain of these microorganisms, the so-called "brown rots, attack the cellulose to the substantial exclusion of the lignin. Others, the white rots, attack both the cellulose and the lignin of wood.

While it is known, therefore, that wood can be altered in composition by the action or microorganisms so as to increase the lignin content thereof, no practical use known to us has resulted from such knowledge. This is probably due to the fact that the action of the microorganisms, as observed and reported in the literature, was too slow for the feasible, commercial exploitation of any process dependent thereon.

While it is also known that wood can be rotted by natural processes involving a combination of separate and distinct processes such as biological and insect-activated processes, the present process has the advantage of being commerciallyieasible by reason of the fact that it gives reproducible results and commercially-interesting yields of lignin within a, comparatively short period of time.

By our present invention, we provide a novel process for conducting the microbiological degradation of cellulose in wood, leaving a ligninenriched product. The improvements in our process reside in the adjustment and control of special conditions of temperature, moisture, in-

oculation and nutrients, which conditions enable up to obtain products containing at least 35 per cent by weight of lignin, within a period not in excess of 90 days. Unless otherwise specifically stated, all statements in this specification and claims dealing with wood compositions refer to wood on the dry basis.

One result of our invention is that wood which is not suitable for direct conversion into vanillin by known processes can be so converted. In view of the commercial importance of vanillin, a widely used flavoring agent, an important practical advantage of this invention is made obvious.

In general, our process involves subjecting wood, suitably moistened and inoculated by means of mycelial fragments, flocs, spores or other pro'pagative entitles, to the degrading microbiological action of certain cellulose-destroying but ligninsparing organisms, at temperatures within the range of about 27 C. to about 32 C., in the presence of certain nutrient substances such as organic sources of nitrogen which accelerate and increase the microbiological action.

We have employed the process of this invention utilizing a number of diflferent woods, and we have found that our process is operable with all the woods employed by us. We have tried hardwoods, and softwoods, woods or various botanical origins, from Gymnosperms and/or Angiosperms, in such forms as wood flour, wood meal, wood chips and sawdust. As examples of specific woods found operableby us may be mentioned pine, hemlock, redwood and birch.

In carrying out our process we first sterilize the wood prior to inoculation. The sterilization can be conducted, for example, by moistening the wood meal or sawdust, or other form of wood employed, with suitable nutrient solutions. Flasks, trays, or other suitable containers are used for the purpose. The containers and contents are placed in a steam pressure sterilizer of suitable capacity and subjected to the eillcient sterilizing action of superheated steam under a pressure of 15 lbs. per square inch for one hour.

We have found that cellulose-destroying but lignin-sparing fungi give the desired results in our-process. Examples of specific fungi which we have tried successfully are Fomitopsis (Fomes) roseus, Fomitopsis (Trametes) subroseus, Lentinus Zepideus, Lenzz'tes saepiaria, Lenez'tes trabea, Polypo'rus palustris, Polyporus schweinitzii, Poria incrassata, Poria microspora, Poria vaillantii (vaporaria) and Poria xantha.

We have found it desirable to use stock cultures of the fungi and carry them on suitable nutrients. A suitable stock culture can be carried on potato-dextrose or potato-maltose agar, prepared either from Difco dehydrated material or from fresh ingredients. A suitable recipe for potato-dextrose or potato-maltose is:

Wash white potatoes and slice with skins on. Put 300 grams in one-half liter of water and boil until soft. Strain through three layers of cheesecloth after washing. Mix the filtrate (about 500 ml.) with 500 ml. of water containing 15 grams of agar already in solution. Make up to one liter. Add 10 grams of dextrose or maltose, and sterilize.

Inocula from the stock cultures of fungi are used to seed the prepared wood. The inocula are applied to the sterlized wood either by chopping up the spawn of vigorous mycelial growth on nutrient agar and distributing the fragments through the wood meal in amounts of about 1 ml. of mycelium to 100 grams of wood or by fragmenting the tufts of mycelium grown in liquid media and adding 5 to ml. of a rich suspension to 50 to 100 grams of wood meal or 5 ml. of bran culture heavily overgrown with mycelium'.

As will be appreciated by those skilled in the art to which this invention pertains, the conditions under which the microbiological process herein is conducted must be carefully controlled in order to obtain the desired results. Important factors which have to be regulated to insure proper functioning of the instant process are temperature, moisture, method of inoculation, pH and the presence of suitable nutrient substances.

The nutrient substances employed in our process are highly important to the success thereof. The substances as used all contain organic nitrogen in greater or lesser amounts together with other compounds of unkown composition which accelerate and stimulate growth. Among the nutrients employed by us are peptone, tryptone, gluten, casein, corn meal, corn steep liquor, wheat bran (whole and water extract), soybean meal, bone meal, fish meal, hoof meal, potato, malt extract and dried yeast.

The nutrients employed and found successful were mixed with water, these solutions or suspensions serving (1) to supply the necessary nutrient substances and (2) to render the wood meal favorably moist for fungus growth.

The nutrients may be employed in varying amounts, based upon the weight of the wood being treated. We have found that the permissive range includes amounts up to 1.2 grams nitrogen per 100 grams of wood. Since the optimum is 0.3 gram nitrogen per 100 grams of wood there is no practical advantage in using greater amounts.

We have found that temperatures within the range of about 27 C. to about 32 C. are required for satisfactory operation. Optimum results are obtained with temperatures of about 28 C. to about 30 C. Temperatures below about 27 C. result in the process being too slow to be feasible; whereas temperatures above 32 C. are too near the upper limit of 35 C. at which temperature growth of the fungi is inhibited.

The moisture is supplied by using the nutrient solutions in amounts of three to one, e. g. 300 ml. of nutrient solution to 100 grams of wood (225 ml. nutrient solution to each 75-gram wood sample).

The permissive pH range at which these fungi operate is pH 4-8, the optimum pH 4-6. Hence the process conditions are such as to give a pH of 4-5 automatically.

The following examples are intended further to illustrate our invention and are not to be construed as limitations thereof.

Example 1 Wood meal (60 mesh) is moistened with peptone solution (6.2 grams peptone per liter of water) in the amounts of 225 ml. of solution to each 75-gram sample of wood meal. The 75- gram sample of nutrient-moistened wood meal is placed as a thin layer on the bottom of a 1-liter Erlenmeyer flask. The flask, plugged with cotton, is then sterlized for one hour in a steam pressure sterilizer with superheated steam at 15 pounds pressure per square inch.

From a vigorous seeding culture of Polyporus palustris growing on wheat bran moistened with peptone solution (in proportions of 70 ml. of solution to 30 grams of bran), 5 ml.' are removed and distributed through the sterile nutrient-moistened wood meal sample. The wood meal sample thus prepared is maintained at a temperature of 28-30 C. for thirty days, giving a. product having a 35 per cent lignin content, and when the conditions were maintained for ninety days microbiological breakdown had been achieved to the extent of giving a product (56.3 per cent lignin) in 84.2 per cent yield (based on lignin).

Example 2 Wood meal is moistened with a nutrient solution (65 grams of hoof meal per liter of water which provides 0.3 gram of nitrogen per grams of wood) in the amounts already given in Example 1. wood meal is placed in a 1-liter Erlenmeyer flask and the flask is plugged and sterilized as in Example 1.

From a vigorous seeding culture of Lentinus Zepideus, grown as in Example 1, the sterlized, nutrient-moistened wood meal sample is inoculated as in Example 1. The wood meal sample thus prepared is maintained at a temperature of 28-30 C. for ninety days, at the end of which time adequate microbiological breakdown has been achieved, giving a product of (53.1 per cent lignin) in 75.4 per cent yield (based on lignin).

Example 3 Example 2 was repeated, using the fungus Polyporus schweinitziz' in place of Lentinus Zepideus for ninety-seven days, giving a product containing 57 per cent lignin in a yield of 87.3 per cent (based on lignin) Example 4 Redwood sawdust of uniform, medium-coarse granular texture is moistened with 10 per cent bran extract solution prepared by boiling 100 grams of commercial wheat bran in one liter of water for one hour, straining through three layers of cheesecloth and adding water to make one liter, in the amounts already given in Example 1. The sample of bran extract moistened redwood sawdust is placed in a l-liter Erlenmeyer flask and the flask pluged and sterilized as in Example 1.

From a vigorous seeding culture of Poria microspora, grown as in Example 1, the sterilized, nutrient-moistened redwood sawdust is inoculated as in Example 1. The redwood sawdust sample thus prepared is maintained at a temperature of 28-30 C. for one hundred and two days, at the end of which time adequate microbiological breakdown has been achieved, giving a product containing 52 per cent lignin.

Example 5 Redwood sawdust of uniform, medium-coarse texture is moistened with a nutrient solution (77 grams white potato waste per liter of water, which provides 0.03 grams of nitrogen per 100 grams of wood) in the amounts already given in Example 1. The methods of preparation, sterilization and inoculation, the temperature range and the length of time, are the same as in Example 4 but the fungus Lenzites trabea is used here. A product containing 49.3 per cent lignin is obtained.

While we have illustrated the practice of our The sample of nutrient-moistened 5 invention in detail, it will be understood by those skilled in the art that minor modifications can be made therein without departing from the spirit of our invention. We intend to cover all such modifications coming within the scope of the apcomprises treating said wood with cellulose-destroying organisms selected from the group consisting of Fomz'topsis (Fo'mes) roseus, Fomz'topsis (Trametes) subroseus, Lentinus Zepideus, Lenzites saepiaria, Lenzites trabea, Polyporus palustris, Polyporus schweinz'tzii, Porz'a incrassata, Poria microspora, Porz'a vaillantii (vaporaria) and Po'ria xantha in the presence of nutrients selected from the group consisting of peptone, tryptone, gluten, casein, com meal, corn steep liquor, wheat bran (whole and water extract), soybean meal, bone meal, fish meal, hoof meal, potato, malt extract and dried yeast at temperatures within the range of about 27 C. to about 32 C. I

2. The process of treating wood so as to form a lignin concentrate having at least 35 per cent by weight lignin content on the dry basis, which comprises sterilizing said wood with steam and then treating said sterilized wood with P012!- porus palustrz's in the presence of nutrients selected from the group consisting of peptone, tryp tone, gluten, casein, corn meal, corn steep liquor, wheat bran (whole and water extract), soybean meal, bone meal, fish meal, hoof meal, potato, malt extract and dried yeast at temperatures within the range of about 28 C. to about 30 C.

3. The process of treating wood so as to form a lignin concentrate having at least 35 per cent by weight lignin content on the dry basis, which 4 comprises sterilizing said wood with steam and then treating said sterilized wood with Lentinus lepideus in the presence of nutrients selected from the group consisting of peptone, tryptone, gluten, casein, corn meal, corn steep liquor, wheat bran (whole and water extract), soybean meal, bone meal, fish meal, hoof me'al, potato, malt extract and dried yeast at temperatures within the range of about 28 C. to about 30 C.

4. The process of treating wood so as to form a lignin'concentrate having at least 35 per cent by weight lignin content on the dry basis, which comprises treating said wood with Pona'microspam in the presence of nutrients selected from the group consisting of peptone, tryptone, gluten, casein, corn meal, corn steep liquor, wheat bran (whole and water extract), soybean meal, bone meal, fish meal, hoof meal, potato, malt extract and dried yeast at temperatures within the range of about 28 C. to about 30 C.

5. The process of claim 1 wherein said wood is sterilized prior to said treatment with cellulosedestroying but lignin-sparing organisms.

ERIC C. KUNZ.

WILLIAM H. WESTON. MARSHALL W. JENNISON. HERMAN R. SWEET. GARRY C. KITCHENS.

References Cited in the file of this patent UNITED STATES PATENTS Number Name Date Re. 22,295 Olson et al. Mar. 30, 1935 1,976,327 Chittenden Oct. 9, 1934 2,431,419 Pearl Nov. 25, 1947 2,440,554 Naghski et al. Apr. 27, 1948 OTHER REFERENCES Virtanen et al.: Suomen Kemistelehti (Biochem. Inst., Helsinki), vol. 193, pages 4-13, 1946. Cited in Chem. Abstracts 1947, cols. 5255-7.

Wiley: Fungi, vol. I, pages 23, 281, 349, 350; vol. II, pages 98, 102, 103, 1947. I 

1. THE PROCESS OF TREATING WOOD SO AS TO FORM A LIGNIN CONCENTRATE HAVING AT LEAST 35 PER CENT BY WEIGHT LIGNIN CONTENT ON THE DRY BASIS, WHICH COMPRISES TREATING SAID WOOD WITH CELLULOSE-DESTROYING ORGANISMS SELECTED FROM THE GROUP CONSISTING OF FOMITOPSIS (FOMES) ROSEUS, FOMITOPSIS (TRAMETES) SUBROSEUS, LENTINUS LEPIDEUS, LENZITES SAEPIARIA, LENZITES TRABEA, POLYPORUS PALUSTRIS, POLYPORUS SCHWEINITZII, (VAPORARIA) AN PORIA MICROSPORA, PORIA VAILLANTII (VAPORARIA) AND PORIA XANTHA IN THE PRESENCE OF NUTRIENTS SELECTED FROM THE GROUP CONSISTING OF PEPTONE, TRYPTONE, GLUTEN, CASEIN, CORN MEAL, CORN STEEP LIQUOR, WHEAT BRAN (WHOLE AND WATER EXTRACT), SOYBEAN MEAL, BONE MEAL, FISH MEAL, HOOF MEAL, POTATO, MALT EXTRACT AND DRIED YEAST AT TEMPERATURES WITHIN THE RANGE OF ABOUT 27* C. TO ABOUT 32* C. 